Glaucoma is defined as the process of ocular tissue destruction caused by a sustained elevation of intra ocular pressure (IOP) above its normal physiological limits1. In open angle glaucoma (OAG), elevated IOP causes a progressive optic neuropathy due to loss of retinal ganglion cells that ultimately leads to blindness2. In angle-closure glaucoma the sudden high rise in IOP often renders the eye blind. Glaucoma is the second leading cause of blindness worldwide3 and the prevalence is increasing worldwide4. Blindness in glaucoma is caused by a degenerative process of the retina and optic nerve, but is functionally associated with impairments in the balance between aqueous humor (AH) secretion and outflow. AH is secreted by cells of the ciliary body (CB) and outflow can occur through one of two pathways: the trabecular meshwork pathway and the uveoscleral pathway5.
Current treatment for glaucoma is not able to restore vision-loss caused by glaucoma, but is focused on IOP reduction6. Controlling IOP has been shown to protect against damage to the optic nerve in glaucoma5, 7. There are five drug classes currently used to achieve IOP reduction: α-adrenergic agonists, β-adrenergic antagonists, cholinergic agonists, prostaglandins and carbon anhydrase inhibitors. If no efficacy in reducing IOP is achieved with any of these drugs, laser therapy can be applied to the trabecular meshwork in order to increase AH outflow. The last therapeutic resource is a surgical procedure to create a new route for AH outflow8.
Current treatments for increased IOP associated with glaucoma have relatively few ocular side effects but may have systemic side effects if the compound reaches the bloodstream9, 10, 11. Treatments that are systemically better tolerated, such as prostaglandins, have many local tolerance issues12. This fact together with the required frequency of instillations in order to maintain adequate levels of IOP makes treatment compliance a challenge for patients13. Failure to comply with therapy cannot only allow for disease progression but can also have a reboot effect causing sudden increases in IOP that can be very damaging to the optic nerve.
Prostaglandins and beta-blockers are the preferred IOP-lowering agents.12,14 Prostaglandins lower IOP extremely well and are safe systemically but have several associated ocular side effects,15 i.e., darkening of the iris color, lash growth, periocular pigmentation, and hyperemia. Less frequent ocular side effects of this drug class are intraocular inflammation, cystoid macular edema, and reactivation of ocular corneal herpes viral infections16. Prostaglandin analogs are contraindicated during pregnancy because of the potential risk of premature labor.
Topical application of beta blockers reduces IOP by decreasing AH production and not by increasing its outflow. Topically administered beta-blockers are absorbed via the conjunctival epithelium, lacrimal channel, nasal mucosa and gastrointestinal tract into the systemic circulation inducing systemic adverse reactions17-19. In the eye, adrenergic receptors have been located at blood vessels that irrigate the ciliary body and trabecular meshwork, where their main effect is vasoconstriction, although their involvement in aqueous humour secretion has also been described. Previous studies in rabbits' eyes showed high density of β-adrenergic receptors in conjunctival, corneal and ciliary process epithelium. β-adrenergic receptors were also present in corneal endothelium, lens epithelium, choroid and extraocular muscle. Most of the β-adrenergic receptors detected in eye belong to the β2-type20-23.
RNA interference (RNAi) is a technology based on the principle that small, specifically designed, chemically synthesized double-stranded RNA fragments can mediate specific messenger RNA (mRNA) degradation in the cytoplasm and hence selectively inhibit the synthesis of specific proteins. This technology has emerged as a very powerful tool to develop new compounds aimed at blocking and/or reducing anomalous activities in defined proteins24,25. Compounds based on RNA interference can be rationally designed to block expression of any target gene, including genes for which traditional small molecule inhibitors cannot be found.26 Examples of successful use of RNAi in therapeutics include inhibition of HIV-1 replication in human cells27 and knock-down of tau and apolipoprotein precursor protein in animal models of Alzheimer's disease.28 Even though RNAi was discovered just over a decade ago, a few of these compounds are already in advanced phases of clinical trials, i.e., RTP801 (Quark Pharmaceuticals, Fremont, Pa., phase II) for treating age-related macular degeneration and ALN-RSV01 (Alnylam Pharmaceuticals, Cambridge, Mass., phase II) for treating respiratory syncytial virus29,30. RNA interference is a very attractive approach to chronic conditions, since upon cessation of treatment the silenced protein has to be re-synthesized in order to recover its biological activity. Hence the effects of compounds based on RNA interference are in general more prolonged than those of conventional treatments24,31.
The eye is a relatively isolated tissue compartment; this particularity provides several advantages to the use of siRNA based therapies. Local delivery of compounds to the eye limits systemic exposure and reduces the amount of compound needed. This allows for local silencing of a gene and reducing the likelihood of wide spread silencing outside the eye. In addition, the immune system has a limited access to the eye; therefore immune responses to the compound are less likely to occur32.
Continuing the work described in WO2006/021817, we have developed an siRNA: SYL040012, as identified in SEQ ID NO: 2, a chemically synthesized, unmodified, 19 bp double-stranded oligonucleotide with dinucleotide overhangs at 3′ of deoxythymidine, able to selectively inhibit synthesis of β2-adrenergic receptor, indicated for the treatment of elevated IOP in patients with ocular hypertension, open angle glaucoma, and other related diseases.
The compound has proven efficacy inhibiting expression of its target in cell cultures and in lowering IOP in normotensive rabbits and in a model of increased IOP in rabbits.